人端粒酶逆转录酶基因和EGFP共表达真核表达载体的构建及鉴定  

Construction and identification of eukaryotic expression vector to co-express hTERT and EGFP

作　　者：代飞[1] 吴军[1] 易绍萱[1] 贺伟峰[1] 陈希炜[1] 张小容[1] 

机构地区：[1]第三军医大学西南医院烧伤研究所,重庆市器官移植基础研究所400038

出　　处：《重庆医学》2004年第8期1206-1208,共3页Chongqing medicine

基　　金：国家自然科学青年基金资助项目 (30 30 0 0 90 ) ;国家重点实验室开放课题基金资助项目

摘　　要：目的　构建并鉴定人端粒酶逆转录酶 (humantelomerasereversetranscriptase ,hTERT)和增强型绿色荧光蛋白(EGFP)双基因共表达重组真核表达质粒。方法　扩增 pGRN1 4 5质粒并酶切回收hTERT片断 ,利用基因重组技术将hTERT连接到 pIRES2 EGFP载体中并酶切鉴定。用NucleofectorTM 技术将构建的重组质粒转染NIH3T3细胞系。结果　成功构建了hTERT和EGFP双基因共表达重组真核表达质粒 ,并成功转染了NIH3T3细胞。结论　重组质粒hTERT pIRES2 EGFP可以被成功导入细胞并表达EGFP 。Objective To construct and identify a recombinant eukaryotic expression plasmid to express both hTERT and EGFP.Methods pGRN145 plasmid was amplified and cut by restriction endonuclease,then the hTERT sequence was collected and recombined with eukaryotic expression vector pIRES2 EGFP by using gene recombination technigue.At last,the recombinant plasmid was identified and transduced into NIH3T3 cells using Nucleofector TM technique.Results The recombinant dicistronic eukaryotic expression plasmid of hTERT and EGFP genes has been constructed and transduced into NIH3T3 cells succesfully.Conclusion The recombinant plasmid hTERT pIRES2 EGFP can be transduced into cells and expressed EGFP successfully.This result a convenient way to detect whether hTERT gene has been transduced into cells,as well as screening positive cells.

关 键 词：HTERT 双基因共表达 PIRES2-EGFP 

分 类 号：Q782[生物学—分子生物学]